Compounds

ABSTRACT

Compounds of general Formula I: ##STR1## are presented which have collagenase inhibition activity and are useful in the management of disease involving tissue degradation and the promotion of wound healing. Representative diseases involving include arthropathy (particularly rheumatoid arthritis), inflammation, dermatological diseases, bone resorption disease and tumor invasion.

This application is a divisional of Ser. No. 07/820,664, filed on Jan.16, 1992, which is a 371 of PCT/GB90/01117 Jan. 16, 1992.

This invention relates to pharmaceutically and veterinarily activeHydroxamic Acid Based Collagenase Inhibitors, which are derivatives ofhydroxamic acid.

The compounds of the present invention act as inhibitors ofmetalloproteases involved in tissue degradation, such as collagenase,which initiates collagen breakdown, stromelysin (protoglycanase),gelatinase and collagenase (IV). There is evidence implicatingcollagenase as one of the key enzymes in the breakdown of articularcartilage and bone in rheumatoid arthritis (Arthritis and Rheumatism,20, 1231-1229, 1977). Potent inhibitors of collagenase and othermetalloproteases involved in tissue degradation are useful in thetreatment of rheumatoid arthritis and related diseases in whichcollagenolytic activity is important. Inhibitors of metalloproteases ifof this type can therefore be used in treating or preventing conditionswhich involve tissue breakdown; they are therefore useful in thetreatment of arthropathy, dermatological conditions, bone resorption,inflammatory diseases and tumour invasion and in the promotion of woundhealing. Specifically, compounds of the present invention may be usefulin the treatment of osteopenias such as osteoporosis, rheumatoidarthritis, osteoarthritis, periodontitis, gingivitis, corneal ulcerationand tumour invasion.

A number of small peptide like compounds which inhibit netalloproteaseshave been described. Perhaps the most notable of these are thoserelating to the angiotensin converting enzyme (ACE) where such agentsact to block the conversion of the decapeptide angiotensin I toangiotensin II a potent pressor substance. Compounds of this type aredescribed in EP-A-0012401.

Certain hydroxamic acids have been suggested as collagenase inhibitorsas in U.S. Pat. No. 4,599,361 and EP-A-0236872. Other hydroxamic acidshave been prepared as ACE inhibitors, for example in U.S. Pat. No.4,105,789, while still others have been described as enkephalinaseinhibitors as in U.S. Pat. No. 4496540.

EP-A-0012401 discloses antihypertensive compounds of the formula:##STR2## wherein R and R⁶ are the same or different and are hydroxy,alkoxy, alkenoxy, dialkylamino alkoxy, acylamino alkoxy, acyloxy alkoxy,aryloxy, alkyloxy, substituted aryloxy or substituted aralkoxy whereinthe substituent is methyl, halo, or nethoxy, amino, alkylamino,dialkylamino, aralkylamino or hydroxyamino;

R¹ is hydrogen, alkyl of from 1 to 20 carbon atoms, including branched,cyclic and unsaturated alkyl groups; substituted alkyl wherein thesubstituent is halo, hydroxy, alkoxy, aryloxy amino, alkylamino,dialkylamino, acrylamino, arylamino, guanidino, imidazolyl, indolyl,mercapto, alkylthio, arylthio, carboxy, carboxamido, carbalkoxy, phenyl,substituted phenyl wherein the substituent is alkyl, alkoxy or halo;aralkyl or heteroaralkyl, aralkenyl or heteroaralkenyl, substitutedaralkyl, substituted heteroaralkyl, substituted aralkenyl or substitutedhetereoaralkenyl, wherein the substituent is halor or dihalo, alkyl,hydroxy, alkoxy, amino, aminomethyl, acrylamino, dialkylamino,alkylamino, carboxyl, haloalkyl, cyano or sulphonamido, aralkyl orhetereoaralkyl substituted on the alkyl portion by amino or acylamino;

R² and R⁷ are hydrogen or alkyl;

R³ is hydrogen, alkyl, phenylalkyl, aminomethylphenylaikyl,hydroxyphenylalkyl, hydroxyalkyl, acetylaminoalkyl, acylaminoalkyl,acylaminoalkyl aminoalkyl, dimethylaminoalkyl, haloalkyl,guanidinoalkyl, imidazolylalkyl, indolylalkyl, mercaptoalkyl andalkylthioalkyl;

R⁴ is hydrogen or alkyl;

R⁵ is hydrogen, alkyl, phenyl, phenylalkyl, hydroxyphenylalkyl,hydroxyalkyl, aminoalkyl, guanidinoalkyl, imidazolylalkyl, indolylalkyl,mercaptoalkyl or alkylthioalkyl;

R⁴ and R⁵ may be connected together to form an alkylene bridge of from 2to 4 carbon atoms, an alkylene bridge of from 2 to 3 carbon atoms andone sulphur atom, an alkylene bridge of from 3 to 4 carbon atomscontaining a double bond or an alkylene bridge as above, substitutedwith hydroxy, alkoxy or alkyl and the pharmaceutically acceptable saltsthereof.

U.S. Pat. No. 4,599,361 discloses compounds of the formula: ##STR3##wherein R¹ is C₁ -C₆ alkyl;

R² is C₁ -C₆ alkyl, benzyl, benzyloxybenzyl, (C₁ -C₆ alkoxy) benzyl orbenzyloxy(C₁ -C₆ alkyl);

a is a chiral centre with optional R or S stereochemistry;

A is a ##STR4## or a --(CR³ ═CR⁴)-- group wherein b and c are chiralcentres with optional R or S stereochemistry;

R³ is hydrogen, C₁ -C₆ alkyl, phenyl or phenyl(C₁ -C₆ alkyl) and R⁴ ishydrogen, C₁ -C₆ alkyl, phenyl(C₁ -C₆ alkyl), cycloalkyl orcycloalkyl(C₁ -C₆ alkyl).

EP-A-0236872 discloses generically compounds of the formula ##STR5##wherein A represents a group of the formula HN(OH)--CO-- orHCO--N(OH)--;

R¹ represents a C₂ -C₅ alkyl group;

R² represents the characterising group of a natural alpha-amino acid inwhich the functional group can be protected, amino groups may beacylated and carboxyl groups can be amidated, with the proviso that R²can not represent hydrogen or a methyl group;

R³ represents hydrogen or an amino, hydroxy, mercapto, C₁ -C₆ alkyl, C₁-C₆ alkoxy, C₁ -C₆ acylamino, C₁ -C₆ -alkylthio, aryl-(C₁ -C₆ alkyl) -,amino-(C₁ -C₆ -alkyl) hydroxy(C₁ -C₆ -alkyl) -, mercaptco(C₁ -C₆ alkyl)or carboxy(C₁ -C₆ alkyl) group, wherein the amino, hydroxy, mercapto orcarboxyl groups can be protected and the amino groups may be acylated orthe carboxyl groups may be amidated;

R⁴ represents hydrogen or a methyl group;

R⁵ represents hydrogen or a C₁ -C₆ acyl, C₁ -C₆ alkoxy-C₁ -C₆ alkyl,di(C₁ -C₆ -alkoxy)methylene, carboxy, (C₁ -C₆ alkyl)carbinyl, (C₁ -C₆alkoxy) carbinyl, arylmethoxy carbinyl, (C₁ -C₆ alkyl)amino carbinyl orarylamino carbinyl group; and

R⁶ represents hydroxy or a methylene group; or

R² and R⁴ together represent a group-(CH₂)_(n) --, wherein n representsa number from 4 to 11; or

R⁴ and R⁵ together represent a trimethylene group;

and pharmaceutically acceptable salts of such compounds, which are acidor basic.

U.S. Pat. No. 4,105,789 generically discloses compounds which have thegeneral formula ##STR6## and salts thereof, wherein R₁ is hydrogen,lower alkyl, phenyl lower alkylene, hydroxy-lower alkylene,hydroxyphenyl lower alkylene, amino-lower alkylene, guanidine loweralkylene, mercapto-lower alkylene, lower alkyl-mercapto-lower alkylene,imidazolyl lower alkylene, indolyl-lower alkylene or carbamoyl loweralkylene;

R₂ is hydrogen or lower alkyl;

R₃ is lower alkyl or phenyl lower alkylene;

R₄ is hydroxy, lower alkoxy or hydroxyamino; and

n is 1 or 2.

U.S. Pat. No. 4,496,540 discloses compounds of the general formula:

    A--B--NHOH

wherein A is one of the aromatic group-containing amino acid residuesL-tryptophyl, D-tryptophyl, L-tyrosyl, D-tyrosyl, L-phenylalanyl, orD-phenylalanyl, and B is one of the amino acids glycine, L-alanine,D-alanine, L-leucine, D-leucine, L-isoleucine, or D-isoleucine; andpharmaceutically acceptable salts thereof.

It would be desirable to improve on the solubility of known collagenaseinhibitors and/or stromelysin inhibitors (whether as the free base orthe salt) and, furthermore, increases in activity have also been sought.It is not a simple matter, however, to predict what variations in knowncompounds would be desirable to increase or even retain activity;certain modifications of known hydroxamic acid derivatives have beenfound to lead to loss of activity.

According to a first aspect of the invention, there is provided acompound of general formula I: ##STR7## wherein: R¹ represents ahydrogen atom or a C₁ -C₆ alkyl, C₁ -C₆ alkenyl, phenyl, phenyl(C₁-C₆)alkyl, C₁ -C₆ alkylthiomethyl, phenylthiomethyl, substitutedphenylthiomethyl, phenyl(C₁ -C₆)alkylthiomethyl orheterocyclylthiomethyl group; or R¹ represents --S--R^(x) wherein R^(x)represents a group ##STR8## R² represents a hydrogen atom or a C₁ -C₆alkyl, C₁ -C₆ alkenyl, phenyl(C₁ -C₆)alkyl, cycloalkyl(C₁ -C₆)alkyl, orcycloalkenyl(C₁ -C₆)alkyl;

R³ represents an amino acid side chain or a C₁ -C₆ alkyl, benzyl, (C₁-C₆)alkoxybenzyl, benzyloxy(C₁ -C₆)alkyl or benzyloxybenzyl group;

R⁴ represents a hydrogen atom or a methyl group;

R⁵ represents a group (CH₂)_(n) A;

n is an integer from 1 to 6; and

A represents a hydroxy, (C₂ -C₇)acyloxy, (C₁ -C₆)alkylthio, phenylthio,(C₂ -C₇)acylamino or N-pyrrolidone group

or a salt and/or N-oxide and/or (where the compound is a thio-compound)a sulphoxide or sulphone thereof.

Hereafter in this specification, the term "compound" includes "salt"unless the context requires otherwise.

As used herein the term "C₁ -C₆ alkyl" refers to a straight or branchedchain alkyl moiety having from one to six carbon atoms, including forexample, methyl, ethyl, propyl, isopropyl, butyl, t-butyl, pentyl andhexyl, and cognate terms (such as "C₁ -C₆ alkoxy") are to be construedaccordingly.

The term "C₁ -C₆ alkenyl" refers to a straight or branched chain alkylmoiety having one to six carbons and having in addition one double bond,of either E or Z stereochemistry where applicable. This term wouldinclude, for example, an alpha, beta-unsaturated methylene, vinyl,1-propenyl, 1- and 2-butenyl and 2-methyl-2-propenyl.

The term "cycloalkyl" refers to a saturated alicyclic moiety having from3 to 8 carbon atoms and includes, for example, cyclopropyl, cyclobutyl,cyclopentyl and cyclohexyl.

The term "cycloalkenyl" refers to an unsaturated alicyclic moiety havingfrom 3 to 8 carbon atoms and includes, for example, cyclopropenyl,cyclobutenyl, cyclopentenyl and cyclohexenyl.

The term "heterocyclylthiomethyl" refers to a methyl group substitutedby a hetrocyclic thiol for example pyridine-2-thiol, pyridine-4-thiol,thiophene-2-thiol or pyrimidine-2-thiol.

The term "substituted", as applied to a phenyl or other aromatic ring,means substituted with up to four substituents each of whichindependently may be C₁ -C₆ alkyl, C₁ -C₆ alkoxy, hydroxy, thiol, C₁ -C₆alkylthiol, amino, halo (including fluoro, chloro, bromo and iodo),triflouromethyl, nitro, --COOH, --COONH₂ or --CONHR^(A), wherein R^(A)represents a C₁ -C₆ alkyl group or the characteristic side chain of anamino acid such as alanine, valine, leucine, isoleucine, phenylalanine,tryptophan, methionine, glycine, serine, threonine, cysteine, tyrosine,asparagine, glutamine, aspartic acid, glutamic acid, lysine, arginine orhistidine.

The term "amino acid side chain" means a characteristic side chainattached to the --CH(NH₂)(COOH) moiety in the following R or S aminoacids: glycine, alanine, valine, leucine, isoleucine, phenylalanine,tyrosine, tryptophan, serine, threonine, cysteine, methionine,asparagine, glutamine, lysine, histidine, arginine, glutamic acid andaspartic acid.

There are several chiral centres in the compounds according to theinvention because of the presence or asymmetric carbon atoms. Thepresence of several asymmetric carbon atoms gives rise to a number ofdiastereomers with the appropriate R or S stereochemistry at each chiralcentre. General formula I and, where appropriate, all other formulae inthis specification are to be understood to include all suchstereoisomers and mixtures (for example racemic mixtures) thereof.Compounds in which the chiral centre adjacent the substituent R³ has Sstereochemistry are preferred.

Further or other preferred compounds include those in which,independently or in any combination:

R¹ represents a hydrogen atom or a C₁ -C₄ alkyl (such as methyl),phenylthiomethyl or heterocyclylthiomethyl (such asthiophenylthiomethyl) group;

R² represents a C₃ -C₆ alkyl (such as isobutyl or n-pentyl) group:

R³ represents a benzyl, 4-(C₁ -C₆)alkoxyphenylmethyl or benzyloxy benzylgroup;

R⁴ represents a hydrogen atom;

n has the value 1, 2 or 3; and/or

A represents a hydroxy, acetoxy, acetylamino, ethylthio or N-pyrrolidonegroup.

Particularly preferred compounds include:

1.4-(N-Hydroxyamino)-2R-isobutylsuccinyl!-L-phenylalanine-N-(2-hydroxyethyl)-amide;

2.4-(N-Hydroxyamino)-2R-isobutylsuccinyl!-L-phenylalanine-N-(2-hydroxyethyl)-N-methylamide;

3. 4-(N-Hydroxyamino)-2R-isobutylsuccinyl!-L-phenylalanine-N-(2-ethylthioethyl) amide;

4.4-(N-Hydroxyamino)-2R-isobutylsuccinyl!-L-phenylalanine-N-(2-N-acetyl-2-aminoethyl)amide;

5.4-(N-Hydroxyamino)-2R-isobutylsuccinyl!-L-phenylalanine-N-(3-(2-pyrrolidone)propyl)amide;

6.4-(N-Hydroxyamino)-2R-isobutylsuccinyl!-L-phenylalanine-N-(3-(2-pyrrolidone)propyl)amide sodium salt;

7.4-(N-Hydroxyamino)-2R-isobutylsuccinyl!-L-phenylalanine-N-(2-acetoxyethyl)amide;

8.4-(N-Hydroxyamino)-2R-isobutyl-3S-methylsuccinyl!-L-phenylalanine-N-(3-(2-pyrrolidone)propyl)amide;

9.4-(N-Hydroxyamino)-2R-isobutyl-3S-methylsuccinyl!-L-phenylalanine-N-methyl-N-(2-hydroxyethyl)amide;

10.4-(N-Hydroxyamino)-2R-isobutyl-3S-methylsuccinyl!-L-phenylalanine-N-(2-hydroxyethyl)amide;

11.4-(N-Hydroxyamino)-2R-isobutyl-3S-methylsuccinyl!-L-phenylaianine-N-(3-(2-pyrrolidone)propyl)amide sodium salt;

and free bases, free acids and salts thereof, where appropriate.Compounds 8 and 5 are especially preferred and compound 8 is the mostpreferred.

Compounds of general formula I may be prepared by any suitable methodknown in the art and/or by the following process, which itself formspart of the invention.

According to a second aspect of the invention, there is provided aprocess for preparing a compound of general formula I as defined above,the process comprising:

(a) deprotecting (for example by hydrogenating) a compound of generalformula III ##STR9## wherein: R¹, R², R³, R⁴ and R⁵ are as defined ingeneral formula I and Z represents a protective group, such as a benzylgroup; or

(b) reacting a compound of general formula IV ##STR10## wherein: R¹, R²,R³, R⁴ and R⁵ are as defined in general formula I,

with hydroxylamine or a salt thereof; and

(c) optionally after step (a) or step (b) converting a compound ofgeneral formula I into another compound of general formula I.

Compounds of general formula I which are sulphoxides or sulphones can bederived from thiol compounds of general formula I by oxidation.Alternatively, thiols of general formula III or IV can be oxidised.

Compounds of general formula I which are disulphide (ie compoundswherein R¹ represents SR^(x)) may be derived from thiol compounds ofgeneral formula I by mild oxidation with, for example, iodine inmethanol.

A compound of general formula III can be obtained by coupling, forexample by conventional coupling techniques, a compound of generalformula IV with an O-protected (for example benzyl) hydroxylamine or byreacting a compound of general formula V ##STR11## wherein: R¹, R² andR³ are as defined in general formula I and Z represents a protectivegroup such as benzyl,

with a compound of general formula VI

    NHR.sup.4 R.sup.5                                          (VI)

A compound of general formula V may be prepared by hydrolysis in thepresence of a base such as sodium hydroxide of a compound of generalformula VII ##STR12## wherein: R¹, R² and R³ are as defined in generalformula I, R⁶ represents a C₁ -C₆ alkoxy, benzyloxy or substituted (eg4-nitro) benzyloxy group, and Z represents a protective group.

A compound of general formula VII may be prepared by coupling, forexample by conventional coupling techniques, a compound of generalformula VIII with an O-protected (for example benzyl) hydroxylamine##STR13## wherein: R¹, R² and R³ are as defined in general formula I andR⁶ represents a C₁ -C₆ alkoxy, benzyloxy or substituted benzyloxy group.

A compound of general formula VIII may be prepared by hydrogenating and(eg thermally) decarboxylating a compound of general formula IX##STR14## wherein: R¹, R² and R³ are as defined in general formula I, R⁸represents a C₁ -C₆ alkyl or benzyl group, R⁶ represents a C₁ -C₆alkoxy, benzyloxy or substituted benzyloxy group and R⁷ represents a C₁-C₆ alkoxycarbonyl or benzyloxycarbonyl group.

A compound of general formula IX may be prepared by reacting asubstituted acid of general formula X ##STR15## wherein: R¹ and R² areas defined in general formula I, R⁸ represents a C₁ -C₆ alkyl or benzylgroup and R⁷ represents a C₁ -C₆ alkoxycarbonyl or benzyloxycarbonylgroup, with

an amino acid derivative of general formula (XI)

wherein: ##STR16## R³ is as defined in general formula I and R⁶represents a C₁ -C₆ alkoxy, benzyloxy or substituted benzyloxy group.

Alternatively, a compound of general formula IV can be prepared byde-esterifying (for example hydrolysing, under acid or base catalysis) acompound of general formula XII ##STR17## wherein: R¹, R², R³, R⁴ and R⁵are as defined in general formula I and R³ represents a C₁ -C₆ alkyl orbenzyl group.

A compound of general formula XII can be prepared in a manner analogousto the preparation of a compound of formula IX by reacting a substitutedacid of general formula XIII ##STR18## wherein: R¹ and R² are as definedin general formula I and R₈ represents a C₁ -C₆ alkyl or benzyl group,

with an amino acid derivative of general formula XIV ##STR19## wherein:R³, R⁴ and R⁵ are as defined in general formula I.

In a further synthetic variant, a compound of general formula X asdefined above wherein R¹ represents a hydrogen atom can be reacted witha compound of general formula XIV to produce a compound of generalformula XV ##STR20## wherein: R², R³, R⁴ and R⁵ are as defined ingeneral formula I, R⁸ represents a C₁ -C₆ alkyl or benzyl group and R⁷represents a C₁ -C₆ alkoxycarbonyl or benzyloxycarbonyl group.

A compound of general formula XV wherein R⁸ represents benzyl and R¹represents benzyloxycarbonyl may be hydrogenated to the malonic acid,then treatment with aqueous formaldehyde and piperidine gives a compoundof formula XVI ##STR21## wherein: R², R³, R⁴ and R⁵ are as defined ingeneral formula I.

Compounds of general formula XVI, by treatment with the appropriatethiols give the acids of general formula IV where R¹ is a substitutedthiomethyl derivative. Thiomethyl derivatives can be oxidised tosulphoxides and sulphones as appropriate.

The starting materials (compounds of general formulae IX, X, XIII andXIV) and reagents described above are either commercially available ormay be produced by conventional processes from commercially availablematerials. For example, when R¹ represents a hydrogen atom, thesubstituted acid of general formula XIII may be prepared by reaction ofan aldehyde XVII

    R.sup.9 CHO                                                (XVII)

wherein R⁹ represents a hydrogen atom or a C₁ -C₅ alkyl C₁ -C₅ alkenyl,phenyl (C₁ -C₅) alkyl, cycloalkyl (C_(-C) ₅) alkyl or cycloalkenyl (C₁-C₅) alkyl group, with a succinate derivative of general formula XVIII,##STR22## wherein: R⁸ represents a C₁ -C₆ alkyl or benzyl group underbase catalysis to give a mixture of acids of general formulae XIXa andXIXb ##STR23## which by hydrogenation, esterification and hydrolysis canbe converted to the acids of the general formula XIII.

Alternatively an ester of general formula XX may be reacted with anester stablised phosphorane of general formula XXI. ##STR24## to yield acompound of general formula XXII ##STR25## wherein R⁸ represents a C₁-C₆ alkyl group, which can be further converted by hydrogenation to theacids of general formula XIII.

In addition the substituted esters may be prepared by reacting an esterof the general formula XXIII ##STR26## wherein Y represents halo and R⁸is as defined above and R¹⁰ is either R¹ or R² as defined above, with amalonate derivative of the general formula XXIV ##STR27## wherein R¹¹ isR² or R¹ as defined above, and the alternative to that substituentemployed in the halo ester.

Compounds of general formulae III and IV are valuable intermediates inthe preparation of compounds of general formula I. According to a thirdaspect of the invention, there is therefore provided a compound ofgeneral formula III. According to a fourth aspect of the invention,there is provided a compound of general formula IV.

As mentioned above, compounds of general formula I are useful in humanor veterinary medicine as they are active inhibitors, ofmetalloproteases involved in tissue degradation.

According to a fifth aspect of the invention, there is provided acompound of general formula I for use in human or veterinary medicine,particularly in the management (by which is meant treatment ofprophylaxis) of disease involving tissue degradation, in particularrheumatoid arthritis, and/or in the promotion of wound healing.

According to a sixth aspect of the invention, there is provided the useof a compound of general formula I in the preparation of an agent forthe management of disease involving tissue degradation, particularlyrheumatoid arthritis, and/or in the promotion of wound healing.Compounds of general formula I can therefore be used in a method oftreating disease involving tissue degradation, particularly rheumatoidarthritis, and/or in a method of promoting wound healing, the method ineither case comprising administering to a human or animal patient aneffective amount of a compound of general formula I.

The potency of compounds of general formula I to act as inhibitors ofcollagenase (a metalloprotease involved in tissue degradation) wasdetermined by the procedure of Cawston and Barrett, (Anal. Biochem., 99,340-345, 1979) and their potency to act as inhibitors of stromelysin wasdetermined using the procedure of Cawston et al (Biochem. J., 195,159-165 1981), both of which techniques are to be described more fullyin the examples and, to the extent that the law allows, are incorporatedby reference herein.

According to a seventh aspect of the invention, there is provided apharmaceutical or veterinary formulation comprising a compound ofgeneral formula I and a pharmaceutically and/or veterinarily acceptablecarrier. One or more compounds of general formula I may be present inassociation with one or more non-toxic pharmaceutically and/orveterinarily acceptible carriers and/or diluents and/or adjuvents and ifdesired other active ingredients.

According to an eighth aspect of the invention, there is provided aprocess for the preparation of a pharmaceutical or veterinaryformulation in accordance with the seventh aspect, the processcomprising admixing a compound of general formula I and apharmaceutically and/or veterinarily acceptable carrier.

Compounds of general formula I may be formulated for administration byany route and would depend on the disease being treated. The may be inthe form of tablets, capsules, powders, granules, lozenges, liquid orgel preparations, such as oral, nasal, topical, or sterile parenteralsolutions or suspensions.

Tablets and capsules for oral administration may be in unit dosepresentation form, and may contain conventional excipients such asbinding agents, for example syrup, acacia, gelatin, sorbitol,tragacanth, or polyvinyl-pyrrollidone; fillers for example lactose,sugar, maize-starch, calcium phosphate, sorbitol or glycine; tabletinglubricant, for example magnesium sterate, talc, polyethylene glycol orsilica; disintegrants, for example potato starch, or acceptable wettingagents such as sodium lauryl sulphate. The tablets may be coatedaccording to methods well known in normal pharmaceutical practice.

Oral liquid preparations may be in the form of, for example, aqueous oroily suspensions, solutions, emulsions, syrups or elixirs, or may bepresented as a dry product for reconstitution with water or othersuitable vehicle before use. Such liquid preparations may containconventional additives such as suspending agents, for example sorbitol,syrup, methyl cellulose, glucose syrup, gelatin, hydrogenated ediblefats; emulsifying agents, for example lecithin, sorbitan monooleate, oracacia: non-aqujeous vehicles (which may include edible oils), forexample almond oil, fractionated coconut oil, oily esters such asglycerine, propylene glycol, or ethyl alcohol; preservatives, forexample methyl or propyl p-hydroxybenzoate or sorbic acid, and ifdesired conventional flavouring or colouring agents.

The dosage unit involved in oral administration may contain from about 1to 250 mg, preferably from about 25 to 250 mg, of a compound of generalformula I. A suitable daily dose for a mammal may vary widely dependingon the condition of the patient and will ultimately depend on thejudgement of the physician or veterinarian. However, a dose of acompound of general formula I of about 0.1 to 300 mg/kg body weight,particularly from about 1 to 100 mg/kg body weight may be appropriate.

For topical application to the skin the drug may be made up into acream, lotion or ointment. Cream or ointment formulations that may beused for the drug are conventional formulations well known in the art,for example, as described in standard text books of pharmaceutics suchas the British Pharmacopoeia.

For topical applications to the eye, the drug may be made up into asolution or suspension in a suitable sterile aqueous or non-aqueousvehicle. Additives, for instance buffers such as sodium metabisulphiteor disodium edeate; preservatives including bactericidal and fungicidalagents, such as phenyl mercuric acetate or nitrate, benzalkoniumchloride or chlorohexidine, and thickening agents such as hypromellosemay also be included.

The dosage employed for the topical administration will, of course,depend on the size of the area being treated. For the eyes each dosewill be typically in the range from 10 to 100 mg of the compound ofgeneral formula I.

The active ingredient may also be administered parenterally in a sterilemedium. The drug depending on the vehicle and concentration used, caneither be suspended or dissolved in the vehicle. Advantageously,adjuvants such as a local anaesthetic, preservative and buffering agentscan be dissolved in the vehicle.

For use in the treatment of rheumatoid arthritis the compounds of thisinvention can be administered by the oral route or by injectionintra-articularly into the affected joint. The daily dosage for a 70 kgmammal will be in the range of 10 mg to 1 gram of a compound of generalformula I.

The following examples illustrate the invention, but are not intended tolimit the scope in any way.

The following abbreviations have been used in the Examples:

DCC--Dicyclohexylcarbodiimide

DCM--Dichloromethane

DCU--Dicyclohexylurea

DIPE--Diisopropyl ether

DMF--N,N-dimethyiformamide

HOBT--Hydroxybenztriazole

NMM--N-Methylmorpholine

TFA--Trifluoroacetic acid

THF--Tetrahydrofuran

WSCDI--N-(Dimethylaminoethyl)-N'-ethylcarbodiimide

EXAMPLES EXAMPLE 14-(N-Hydroxyamino)-2R-isobutylsuccinyl!-L-phenylalanine-N-(2-hydroxyethyl)-amide##STR28## (a)4-Benzyloxy-3-benzyloxycarbonyl-2R-isobutylsuccinyl!-L-phenylalaninemethyl ester

Benzyl (2-benzyloxycarbonyl-5-methyl-3R-tert-butoxy-carbonyl)-hexanoate(52 g, 115 mmol) was stirred at room temperature with 5% water in TFA(250 ml) for 1.5 h. After this time the TFA was evaporated under reducedpressure then the residue was azeotroped with toluene (3×250 ml).

The crude acid from this reaction was dissolved in DCM/DMF (4:1), thenHOBT (16 g, 118 mmol), NMM (12 g, 118 mmol) and WSCDI (22 g, 115 mmol)were added at room temperature. After 20 minutes a further equivalent ofNMM (12 g, 118 mmol) was added followed by L-phenylalanine methyl esterhydrochloride (23 g, 107 mmol). This solution was stirred overnight andthen concentrated under vacuum. The oily residue was dissolved in DCMthen washed with 10% citric acid (2×250 ml), with 10% sodium bicarbonate(2×250 ml) and once with saturated brine (250 ml). The organic layer wasdried (sodium sulphate), filtered then the solvent removed under reducedpressure to give the title compound as an oil (50.9 g, 79%).

δ_(H) (250 MHz, CDCl₃, 3:1 mixture of diastereomers) Major diastereomer0.72 (3H, d, J=6 Hz, CH(CH₃)₂), 0.74 (3H, d, J=6 Hz, CH(CH₃)₂),0.80-1.00 (2H, m, CHCH₂ +CHMe₂), 1.40-1.60 (2H, m, CHCH₂ +CHCH₂), 2.95(1H, dd, J=14,6 Hz, CH₂ Ph), 3.07 (1H, dd, J=14,5 Hz, CH₂ Ph) , 3.64(3H, s, CO₂ Me) , 3.82 (1H, d, J=10 Hz, CH(CO₂ Bn)₂), 4.82 (1H, m,CHCO), 5.0-5.2 (2H, m, OCH₂ Ph), 6.2 (1H, d, J=8 Hz, NH), and 7.10-7.40(15H, m, Ph). Minor diastereomer shows 0.63 (3H, d, J=6 Hz, CH(CH₃)₂) ,0.68 (3H, d, J=6 Hz, CH(CH₃)₂), 3.67 (3H, s, CO₂ Me), and 3.75 (1H, d,J=8 Hz, CH(CO₂ Bn)₂)

(b) 4-Hydroxy-2R-isobutylsuccinyl!-L-phenylalanine methyl ester

The product from above (50.9 g, 91 mmol) was dissolved in ethanol (100ml) and stirred at room temperature with activated charcoal pellets for1 h. 10% Palladium on charcoal (20 g) in ethyl acetate was slurried intothe ethanolic solution. Cyclohexene (20 ml) in ethanol (100 ml) wasadded and the mixture was brought to reflux for 5 h. The reactionmixture was filtered to remove the catalyst, then the solventsevaporated under reduced pressure to leave a yellow oil (29.8 g). Thisoil was taken up in xylene (500 ml) and heated at reflux for 10 minutes.The xylene was removed under reduced pressure to leave the crudematerial as an oil (26.5 g).

(c) 4-(N-Benzyloxyamino)-2R-isobutylsuccinyl!-L-phenylalanine methylester

The crude acid (26.5 g, 79 mmol) was dissolved in DCM/DMF (4:1, 500 ml),then NMM (9.6 g, 95 mmol), HOBT (12.8 g 95 mmol) and WSCDI (18.2 g, 95mmol) added and the mixture stirred at room temperature until tlcindicated complete conversion to the activated ester (about 10 minutes).To this solution containing the active ester was addedbenzylhydroxylamine hydrochloride (15.2 g, 95 mmol) and a furtherequivalent of NMM (9.6 g, 95 mmol) in the solvent mixture (80 ml). Afterstirring at room temperature overnight DCM (250 ml) was added then themixture washed with citric acid (2×250 ml), 10% sodium bicarbonatesolution (2×250 ml) and brine (250 ml) then finally dried over sodiumsulphate. The solution was filtered and the solvent removed underreduced pressure to give an oil (27.2 g) which was purified by columnchromatography using ether as an eluant to give the title compound (11g, 23.7 mmol, 30%).

δ_(H) (250 MHz, CDCl₃) 0.84 (6H, m, CH(CH₃)₂), 1.16 (1H, m, CHMe₂), 1.51(2H, m, CH₂ CHMe₂), 2.1-2.4 (2H, bm, CH₂ CONHOBn), 2.73 (1H, m, CH₂CHCO), 3.06 (2H, d, J=6 Hz, CH₂ Ph), 3.68 (3H, s, CO₂ Me), 4.8-5.0 (3H,s+m, OCH₂ Ph and COCHNH), 6.25 (1H, d, J=8 Hz, NH), 7.05-7.50 (10H, m,Ph), and 8.66 (1H, s, NHOBn).

(d) 4-(N-Benzyloxyamino)-2R-isobutylsuccinyl!-L-phenylalanine

4-(N-Benzyloxyamino)-2R-isobutylsuccinyl!-L-phenylalanine methyl ester(9.5 g, 21 mmol) was dissolved in methanol (200 ml) and lithiumhydroxide solution (0.5N, 84 ml, 42 mmol) was added with stirring atroom temperature. When the reaction was complete, as judged from tlc,the methanol was removed by evaporation and the remaining aqueous phasewas acidified to pH1 with citric acid. The precipitated solid wasfiltered off and dried, while the fitrate was extracted with DCM (500ml) and dried over sodium sulphate. Solvent removal from the organicphase left an oil (5.38 g) which could be recrystallised fromdiisopropyl ether and methanol to give material which was identical withthe solid which precipitated during acidification. These two batcheswere combined to give the title compound (6.40 g, 15 mmol, 71%)

m.p. 161-162° C.

nu_(max) (KBr) 3300, 3020, 2980, 1710, 1650, 1630, 1550, 1265, 740, and700 cm⁻¹

δ_(H) (250 MHz, CDCl₃ /D₆ -DMSO) 0.80-0.87 (7H, m), 1.50 (2H, bm),2.0-2.1 (2H, m), 2.91-3.14 (2H, m, CH₂ Ph), 4.77 (2H, s, OCH₂ Ph), and7.18-7.36 (10H, m, Ph).

δ_(C) (62.9 MHz, D₆ -DMSO) 174.1, 173.1, 167.7, 137.9, 129.2-126.4,76.9, 53.3, 40.7, 39.9, 36.8, 35.8, 25.3, 23.5, and 22.1

(e)4-(N-Hydroxyamino)-2R-isobutylsuccinyl!-L-phenylalanine-N-(2-hydroxyethyl)-amide

4-(N-Benzyioxyamino)-2R-isobutylsuccinyl!-L-phenylalanine (7.50 g, 17.6mmol) was dissolved in DCM (100 ml) and cooled in ice. Triethylamine(1.96 g, 19.4 mmol) was added together with ethylchloroformate (2.10 g,19.4 mmol) and after 10 minutes ethanolamine (1.55 g, 21.1 mmol) in DCH(10 ml) was added. After 3 h at room temperature the reaction mixturewas diluted with ethyl acetate then washed with sodium bicarbonatesolution and brine, and finally dried over sodium sulphate. Solventremoval under reduced pressure gave the crude benzyl hydroxamate whichwas recrystallised from ethyl acetate/hexane (2.6 g, 5.5 mmol)

The crude material from above was dissolved in cyclohexene/ethanol (10%solution, 55 ml), 10% palladium on charcoal (250 mg) was added then themixture refluxed until starting material had dissappeared by tlc (ca. 30minutes). The catalyst was removed by filtration, and the solventremoved under reduced pressure to leave a solid which could berecrystallised from methanol and DIPE. The required product (1.54 g,4.00 mmol, 74%) was collected by filtration.

m.p. 156-158° C.

α!_(D) =-21.5 (c=1, MeOH)

nu_(max) (KBr) 3300, 2950, 1650, 1550, and 700 cm⁻¹

δ_(H) (250 MHz, CDCl₃) 0.72 (3H, d, J=6 Hz, CH(CH₃)₂), 0.77 (3H, d, J=6Hz, CH(CH₃)₂), 0.95 (1H, m, CHCH₂), 1.28 (2H, m, CH(CH₃)₂ +CHCH₂), 1.92(2H, m, CH₂ CONHOH), 2.61 (1H, bm, CHCO), 2.80 (1H, dd, J=14,12 Hz, CH₂Ph), 3.00-3.20 (3H, m, NCH₂ +CH₂ Ph), 4.41 (1H, m, NCHCO), 4.65 (1H, bt,OH), 7.22 (5H, m, Ph), 7.86 (1H, t, J=6 Hz, CONHCH₂), 8.07 (1H, d, J=8Hz, CONH), and (8.76 (1H, s, NHOH).

δ_(C) (62.9 MHz, D₆ -DMSO) 174.0, 171.2, 138.3, 129.2, 128.1, 126.2,59.8, 54.0, 41.6, 37.3, 35.8, 25.3, 23.5, and 22.0.

Analysis calculated for C₁₉ H₂₉ N₃ O₅ Requires C 60.14H 7.70N 11.07Found C 59.97H 7.68N 11.10

EXAMPLE 24-(N-Hydroxyamino)-2R-isabutylsuccinyl!-L-phenylalanine-N-(2-hydroxyethyl)-N-methylamide.##STR29##

Using the procedure described in Example 1e4-(N-Benzyloxyamino)-2R-isobutylsuccinyl!-L-phenylalanine (0.5 g, 1.17mmol ) was coupled with N-methylethanolamine (100 mg, 1.29 mmol) thenthe product hydrogenated to give the title compound (97 mg, 0.25 mmol,21%)

m.p. 136.0-137.0° C.

α!_(D) =+1.1 (c=1, MeOH)

nu_(max) (KBr) 3600-3100, 2960, 1680, 1560, 940, 750, and 700 cm⁻¹

δ_(H) (250 MHz, CDCl₃ /D₆ -DMSO, 1:1) 0.74 (6H, m, CH(CH₃)₂), 1.00 (1H,m, CH₂ CHMe₂), 1.36 (2H, m, CH₂ CHMe₂), 2.00 (2H, m, CH₂ NHOH),2.60-3.00 (3H, m, +3H, 2×s), 3.39 (2H, m), 4.42-4.70 (1H, m, OH),4.80-5.00 (1H, m, CH₂ Ph), 7.14 (5H, m, Ph), 7.97 (1H, d, J=7 Hz, NH),and 8.50 (1H, s, NHOH)

δ_(C) (62.9 MHz, D₆ -DMSO, 1:1 Mixture of Rotamers) 173.9, 173.8, 171.3,170.3, 167.5, 138.2, 137.9, 129.4, 128.1, 126.4, 126.3, 58.7, 58.4,51.2, 50.2, 50.0, 49.7, 40.7, 40.5, 39.8, 37.7, 37.4, 36.0, 35.9, 33.9,25.4, 23.5, 23.4, and 22.1.

EXAMPLE 34-(N-Hydroxyamino)-2R-isobutylsuccinyl!-L-phenylalanine-N-(2-ethylthioethyl) amide. ##STR30##

Using the procedure described in Example 1e4-(N-Benzyloxyamino)-2R-isobutylsuccinyl!-L-phenylalanine (0.5 g, 1. 17mmol) was coupled with 2-(thioethyl)-1-aminoethane hydrochloride (183mg, 1.29 mmol) then the product hydrogenated to give the title compound(163 mg).

m.p. 169-171° C.

α!_(D) =-19.7° (c=1, MeOH)

nu_(max) (KBr) 3280, 2950, 2920, 1655, 1640, and 1540 cm⁻¹

δ_(H) (250 MHz, D₆ -DMSO) 0.72 (3H, d, J=6 Hz), 0.77 (3H, d, J=7 Hz),0.95 (1H, m), 1.17 (3H, t, J=7 Hz), 1.29 (2H, m), 1.93 (2H, m), 2.49(4H, m), 2.62 (1H, m), 2.82 (1H, dd, J=14,10 Hz), 3.03 (1H, dd, J=14,5Hz), 3.19 (2H, m), 4.40 (1H, m), 7.22 (5H, m), 8.07 (2H, m), 8.75 (1H,s), and 10.39 (1H, s).

δ_(C) (62.9 MHz, D₆ -DMSO) 174.0, 171.1, 167.7, 138.3, 129.2, 128.1,126.3, 54.1, 40.8, 40.7, 38.8, 37.3, 35.8, 30.0, 25.3, 24.9, 23.5, 22.0,and 14.9.

EXAMPLE 44-(N-Hydroxyamino)-2R-isobutylsuccinyl!-L-phenylalanine-N-(2-N-acetylethyl)amide. ##STR31##

Using the procedure described in Example 1e4-(N-Benzyloxyamino)-2R-isobutylsuccinyl!-L-phenylalanine (0.43 g, 1.0mmol) was coupled with N-acetyl-1,3-ethyldiamine (133 mg, 1.30 mmol)then the product hydrogenated to give the title compound (176 mg, 0.42mmol, 42%)

m.p. 167-169° C.

α!_(D) =-4.2° (c=1, MeOH)

nu_(max) (KBr) 3280, 2930, 1640, 1540, and 700 cm⁻¹

δ_(H) (250 MHz, D₆ -DMSO) 0.72 (3H, d, J=6 Hz), 0.77 (3H, d, J=6 Hz),1.30 (2H, t, J=10 Hz), 1.78 (3H, s), 2.06 (1H, m), 2.18 (1H, m), 2.87(2H, m), 3.04 (2H, s), 3.13 (2H, q, J=6 Hz), 3.36 (1H, m), 7.21 (5H, m),7.78 (1H, s), 7.98 (1H, s), 8.08 (1H, d, J=8 Hz) and 8.77 (1H, s).

δ_(C) (62.9 MHz, D₆ -DMSO) 174.8, 171.2, 169.4, 167.8, 138.3, 129.2,128.1, 128.2, 54.2, 37.2, 35.8, 25.3, 23.4, 22.8 and 22.01.

Analysis calculated for C₂₁ H₃₂ N₄ O₅.0.4H₂ O Required C 58.97H 7.73N13.10 Found C 59.07H 7.60N 12.90

EXAMPLE 54-(N-Hydroxyamino)-2R-isobutylsuccinyl!-L-phenylalanine-N-(3-(2-pyrrolidone)propyl)amide. ##STR32##

Using the procedure described in Example 1e4-(N-Benzyloxyamino)-2R-isobutylsuccinyl!-L-phenylalanine (0.43 g, 1.0mmol) was coupled with 1-(3-aminopropyl)-2-pyrrolidinone (180 mg, 1.26mmol) then the product hydrogenated to give the title compound (280 mg,0.61 mmol, 61%)

m.p. 174-176° C.

α!_(D) =-8.7° (c=1.35, MeOH)

nu_(max) (KBr) 3270, 3220, 2960, 1660, 1640, and 1525 cm⁻¹

δ_(H) (250 MHz, D₆ -DMSO) 0.72 (3H, d, J=6 Hz), 0.77 (3H, d, J=6 Hz),0.98 (1H, m), 1.33 (2H, m) , 1.52 (2H, m), 1.87-2.06 (4H, m), 2.20 (2H,t, J=8 Hz), 2.62 (1H, m), 2.83 (1H, dd, J=14,9 Hz), 2.99 (1H, t, J=8Hz), 3.10 (2H, t, J=7 Hz), 3.28 (2H, m), 4.39 (1H, m), 7.22 (5H, m),7.91 (1H, m), 8.09 (1H, d, J=8 Hz), 8.80 (1H, bs), and 10.4 (1H,bs).

δ_(C) (62.9 MHz, D₆ -DMSO) 174.0, 171.0, 167.7, 138.3, 129.2, 128.1,126.2, 54.2, 46.6, 40.9, 40.6, 39.6, 38.7, 37.4, 36.5, 35.3, 30.6, 26.9,25.3, 23.4, 22.1 and 17.7.

Analysis calculated for C₂₄ H₃₆ N₄ O₅ Required C 62.59H 7.88N 12.16Found C 62.67H 7.96N 12.22

EXAMPLE 64-(N-Hydroxyamino)-2R-isobutylsuccinyl!-L-phenylalanine-N-(3-(2-pyrrolidone)propyl)amide sodium salt. ##STR33##

4-(N-Hydroxyamino)-2R-isobutylsuccinyl!-L-phenylalanine-N-(3-(1-pyrrolidone)propyl)amide (50 mg, 0.109 mmol) was dissolved in methanol (20 ml) and sodiumhydroxide solution (1.0M, 0.11 ml) added to give a homogeneous solution.The methanol was removed under reduced pressure then the residualaqueous solution freeze dried to give the title compound (52 mg, 0.108mmol, 99%).

δ_(H) (250 MHz, D₆ -DMSO) 0.66 (3H, d, J=6 Hz) , 0.75 (3H, d, J=6 Hz),0.94 (1H, m) , 1.04 (2H, m) , 1.56 (2H, m), 1.92 (3H, m), 2.08 (1H, dd,J=14,8 Hz), 2.14 (2H, t, J=8 Hz), 2.45 (1H, m), 2.83 (1H, dd, J=14,10Hz), 3.03 (2H, d, J=6 Hz), 3.13 (4H, m), 3.23-3.48 (6H, m), 4.35 (1H,m), 7.20 (5H, m), 8.20 (1H, d, J=8 Hz), and 8.53 (1H, s).

EXAMPLE 74-(N-Hydroxyamino)-2R-isobutylsuccinyl!-L-phenylalanine-N-(2-acetoxyethyl)amide. ##STR34##

4-(N-Hydroxyamino)-2R-isobutylsuccinyl!-L-phenylalanine-N-(2-hydroxyethyl)-amide(Example 1e, 148 mg, 0.32 mmol) was mixed with dimethylamino pyridine(40 mg, 0.33 mmol) in DCM at -30°, then acetic anhydride (32 mg, 0.32mmol) was added and the reaction stirred for 25 min. The mixture waspartitioned between ethyl acetate and water, the organic layer separatedand washed sequentially with sodium bicarbonate, citric acid and brinethen dried over sodium sulphate. Purification by column chromatography(ethyl acetate as eluant) gave protected material (130 mg) which washydrogenated as before to give the title compound (62 mg, 0.15 mmol,46%).

m.p. 136-137° C.

α!_(D) =-19.9° (c=1.2, MeOH)

nu_(max) (KBr) 3230, 2955, 1745, 1660, 1645, 1550 and 1235 cm⁻¹

δ_(H) (250 MHz , CDCl₃ /D₆ -DMSO) 0.71 (3H, d, J=6 Hz), 0.75 (3H, d, J=6Hz), 0.99 (1H, m), 1.2-1.4 (2H, m), 1.96 (1H, m+3H, s), 2.11 (1H, dd,J=14,8 Hz), 2.59 (1H,m), 2.86 (1H, dd, J=14,9 Hz), 3.06 (1H, dd, J=14,5Hz), 3.30 (2H, m), 3.95 (2H, t, J=6 Hz) , 4.44 (1H, m), 7.16 (5H, m),and 8.00 (2H, m).

δ_(C) (62.9 MHz , D₆ -DMSO) 174.1, 171.3, 170.1, 168.0, 138.0, 129.1,127.9, 126.0, 78.8, 62.4, 41.0, 40.9, 37.8, 37.3, 35.8, 25.3, 23.1, 21.9and 20.7.

Analysis calculated for C₂₁ H₃₁ N₃ O₆.0.4H₂ O Required C 58.84H 7.48N9.80 Found C 58.91H 7.33N 9.55

EXAMPLE 84-(N-Hydroxyamino)-2R-isobutyl-3S-methylsuccinyl!-L-phenylalanine-N-(3-(2-pyrrolidone)propyl)amide. ##STR35## (a)2S-tert-Butyl(3,3-di(benzyloxycarbonyl)-2,5-dimethyl)hexanoate

Benzyl (2-benzyloxycarbonyl-5-methyl) pentanoate (100 g, 0.29 mol) wasdissolved in dry DMF (150 ml) and cooled while potassium tert-butoxide(31.1 g, 0.28 mol) was added portionwise over 10 minutes. This was thenstirred for a further 1 hour until the solid had dissolved. To theresultant mixture, cooled to -20 to -30°, was added tert-butyl2S-bromopropionate (60.6 g, 0.29 mol) in dry DMF (50 ml) over about 30minutes. The reaction mixture was left at -20° for 3 days then 5° for 1day before working up by partitioning between ether and ammoniumchloride solution. The aqueous layer was extracted three times withether then the combined organic layers washed with brine and dried.Solvent removal gave the crude title compound (141.2 g, 0.30 mol, 100%)

α!_(D) =33.0° (c=1.00, MeOH)

δ_(H) (250 MHz , CDCl₃) 0.82 (6H, d, J=6 Hz), 1.28 (3H, t, J=7 Hz), 1.40(9H, s), 1.46 (2H, m), 1.76 (1H, m), 1.92 (1H, dd, J=6,2 Hz), 5.11 (4H,m), and 7.27 (10H, m)

δ_(C) (62.9 MHz, CDCl₃) 172.7, 170.3, 135.3, 128.2, 80.8, 66.9, 58.7,45.2, 42.5, 27.8, 24.4, 23.9, 23.8, and 14.3.

(b) 2S,3R tert-Butyl (3-carboxylic acid-2,5-dimethyl)-hexanoate

The crude material from Example 13a (141.2 g, 0.30 mol) was taken up nethanol (200 ml) and refluxed with activated charcoal (10 g) for 1 hourto remove catalyst poisons. Cyclohexene (100 ml) and 10% palladium oncharcoal (14 g) was added and the mixture refluxed for 2 hours. Thecatalyst was removed by filtration through celite and the solventremoved under reduced pressure.

The residual oil was taken up in xylene (200 ml) and refluxed for 30minutes to effect decarboxylation. The solution was extracted withsodium carbonate solution (3×300 ml) this aqueous solution washed withether then acidified to pH 5 with citric acid. The acidic solution wasextracted with ethyl acetate (3×200 ml) then dried over sodium sulphate.Solvent removal then gave the crude title compound (53.2 g, 0.22 mol,73%)

α!_(D) =8.0° (c=1.00, MeOH)

δ_(H) (250 MHz, CDCl₃) 0.76 (6H, 2×d, J=6 Hz), 1.02 (3H, m), 1.28 (9H,s), 1.46 (2H, m), 2.42 (1H, m), and 2.58 (1H, m)

δ_(C) (62.9 MHz, CDCl₃) 173.9, 128.7, 46.5, 43.0, 39.1, 27.3, 26.2,23.5, 21.4, and 14.9.

(c) 4-(tert-Butyloxy)-2R-isobutyl-3S-methylsuccinyl!-L-phenylalaninemethyl ester

The crude acid from Example 13b (45.0 g, 0.18 mol) was dissolved in DCM,then HOBT (24.9 g, 0.18 mol) added. The solution was cooled and NMM (18g, 0.18 mol), phenylalanine methyl ester hydrochloride (36.1 g, 0.17mol) and DCC (38 g, 0.18 mol) were added. This solution was stirredovernight, concentrated under vacuum then the precipitated DCU filteredoff. The oily residue was dissolved in ethyl acetate then washed with10% citric acid (2×250 ml), with 10% sodium bicarbonate (2×250 ml) andonce with saturated brine (250 ml). The organic layer was dried (sodiumsulphate), filtered then the solvent removed under reduced pressure togive the crude title compound as an oil (80.6 g, 0.20 mol, 120%).

(d) 4-(N-Benzyloxyamino)-2R-isobutyl-3S-methyl-succinyl!-L-phenylalaninemethyl ester

The crude tert-butyl ester (80.6 g, 0.20 mol) was dissolved intrifluroacetic acid/water (95:5, 85 ml) and left at 4° C. overnight. Thesolution was taken up in DCM, the aqueous layer re-extracted with DCMthen the combined organic layers extracted with sodium bicarbonate (5×50ml). The basic layer was acidified to pH 4 with citric acid thenextracted with ethyl acetate. Drying and solvent removal gave therelated acid (40.4 g, 0.115 mol. 69%).

The crude acid (40.4 g, 115 mmol) was dissolved in DCM/DMF (4:1, 500ml), then HOBT (17.18 g 127 mol) and DCC (26.1 g, 127 mmol) were addedand the mixture stirred at room temperature until tlc indicated completeconversion to the activated ester (about 10 minutes). To this solutioncontaining the active ester was added benzylhydroxylamine (15.6 g, 127mmol). After stirring at room temperature overnight DCM was removedunder vacuum, the residue taken up in ethyl acetate then precipitatedDCU removed by filtration. The solution was washed with citric acid(2×250 ml), 10% sodium bicarbonate solution (2×250 ml) and brine (250ml) then finally dried over sodium sulphate. The solvent was removedunder reduced pressure to give an oil (45.6 g) which was purified byrecrystallisation from ethanol and DIPE (7.66 g, 17 mmol, 15%).

δ_(H) (250 MHz , CDCl₃) 0.47 (3H, d, J=7 Hz), 0.74 (3H, d, J=6 Hz), 0.83(3H, d, J=6 Hz), 1.35 (2H, m), 1.94 (1H, dd, J=7,11 Hz), 2.38 (1H, m),2.83 (1H, dd, J=14,11 Hz), 3.06 (1H, dd, J=5,14 Hz), 3.29 (3H, s), 3.62(3H, s), 4.58 (1H, m), 4.77 (2H, s), 7.20 (5H, m), 7.38 (5H, s), and8.49 (1H, d, J=8 Hz).

(e) 4-(N-Benzyloxyamino)-2R-isobutyl-3S-methyl-succinyl!-L-phenylalanine

4-(N-Benzyloxyamino)-2R-isobutyl-3S-methyl-succinyl!-L-phenylalaninemethyl ester (7.66 g, 17 mmol) was dissolved in methanol (120 ml) andsodium hydroxide solution (1.0M, 20.3 ml, 20.3 mmol) was added withstirring at room temperature. When the reaction was complete, as judgedfrom tlc, the methanol was removed by evaporation the residue extractedwith ether to remove starting material (2.1 g, 4.6 mmol, 21% recovered).The aqueous phase was acidified to pH4 with citric acid and extractedwith ethyl acetate to give the title compound (5.88 g, 13.3 mmol, 79%).

δ_(H) (250 MHz, CDCl₃) 0.47 (3H, d, J=7 Hz), 0.74 (3H, d, J=6 Hz), 0.83(3H, d, J=6 Hz), 1.35 (2H, m), 1.94 (1H, dd, J=7,11 Hz), 2.38 (1H, m),2.83 (1H, dd, J=14,11 Hz), 3.06 (1H, dd, J=5,14 Hz), 3.29 (3H, s), 3.62(3H, s), 4.58 (1H, m), 4.77 (2H, S), 7.20 (5H, m), 7.38 (5H, s), and8.49 (1H, d, J=8 Hz).

(f)4-(N-Hydroxyamino)-2R-isobutyl-3S-methyl-succinyl!-L-phenylalanine-N-(3-(2-pyrrolidone)propyl)amide.

4-(N-Benzyloxyamino)-2R-isobutyl-3S-methylsuccinyl!-L-phenyl alanine(400 mg, 0.9 mmol), HOBT (134 mg, 1.0 mmol) and NMM (102 mg, 1,0 mmol)were dissolved in DCM/DMF (4:1, 10 ml) and cooled in ice.1-(3-Aminopropyl)-2-pyrrolidinone (142 mg, 1.0 mmol) was added togetherwith WSCDI (192 mg, 1.0 mmol). After 2 h at room temperature thereaction mixture was diluted with ethyl acetate then washed with sodiumbicarbonate solution and brine, then dried over sodium sulphate. Solventremoval under reduced pressure gave the crude benzyl hydroxamate whichwas recrystallised from ethyl acetate/hexane (385 mg, 0.68 mmol)

The material from above was dissolved in cyclohexene/ethanol (10%solution, 20 ml), 10% palladium on charcoal (50 mg) was added then themixture refluxed until starting material had dissappeared by tlc (ca. 30minutes). The catalyst was removed by filtration, and the solventremoved under reduced pressure to leave a solid which could berecrystallised from ethyl acetate/ethanol. The required product (243 mg,0.51 mmol, 76%) was collected by filtration.

m.p. 198-200° C.

α!_(D) =73.0° (c=1.00, MeOH)

nu_(max) (KBr) 3380, 2960, 1635, 1345, 1030, and 715 cm⁻¹

δ_(H) (250 MHz , D₆ -DMSO) 0.44 (3H, d, J=7 Hz, CHCH₃), 0.72 (3H, d, J=6Hz, CH(CH₃)₂), 0.80 (3H, d, J=6 Hz, CH(CH₃)₂), 1.32 (2H, m), 1.53 (2H,q, J=7 Hz), 1.92 (4H, m), 2.21 (2H, t, J=8 Hz), 2.37 (1H, m), 2.79 (1H,m), 2.93 (1H, d, J=5 Hz), 3.01 (2H, m), 3.13 (2H, t, J=7 Hz), 3.32 (6H,m), 4.54 (1H, m), 7.25 (5H, m), 7.80 (1H, t, J=7 Hz), 8.20 (1H, d, J=8Hz, NH), 8.69 (1H, s), and 10.36 (1H, s).

δ_(C) (62.9 MHz , D₆ -DMSO) 174.0, 173.9, 171.4, 171.2, 138.2, 129.3,128.0, 126.3, 54.6, 47.0, 46.7, 37.7, 36.3, 30.7, 27.3, 25.4, 24.3,21.5, 17.7 and 16.1.

Analysis calculated for C₂₅ H₃₈ N₄ O₅.0.5 H₂ O Required C 62.20H 7.93N11.50 Found C 63.27H 8.07N 11.50

EXAMPLE 94-(N-Hydroxyamino)-2R-isobutyl-3S-methylsuccinyl!-L-phenylalanine-N-methyl-N-(2-hydroxyethyl)amide. ##STR36##

Using the procedure described in Example 13f4-(N-Benzyloxyamino)-2R-isobutyl-3S-methylsuccinyl!-L-phenylalanine (0.4g, 0.91 mmol) was coupled with N-methylethanolamine (75 mg, 1.00 mmol)then the product hydrogenated to give the title compound (100 mg, 0.25mmol, 27%)

m.p. 182-183° C.

α!_(D) =27.0° (c=0.25, MeOH)

nu_(max) (KBr) 3400, 3240, 2960, 1660, and 1545 cm⁻¹

δ_(H) (250 MHz , D₆ -DMSO, mixture of rotamers) 0.46+0.54 (3H, d, J=6Hz, CHCH₃), 0.72+0.79 (6H, m, CH(CH₃)₂), 1.3 (2H, m), 2.0 (1H, m), 2.4(1H, m), 2.81+3.03 (3H, s, N(CH₃)), 2.8-3.9 (8H, m), 5.0 (1H,m), 7.27(5H, m), 8.35 (1H, d, J=8 Hz, NH), and 8.73 (1H,m).

δ_(C) (62.9 MHz, D₆ -DMSO, mixture of rotamers), 173.3, 173.2, 171.6,171.4, 171.0, 138.3, 138.0, 129.4, 128.1, 128.0, 126.4, 126.3, 58.8,58.5, 51.4, 50.2, 50.1, 49.9, 46.7, 40.1, 37.4, 37.1, 36.2, 33.9, 25.5,25.4, 24.2, 24.1, 21.8, 21.7, 16.3, and 16.2.

Analysis calculated for C₂₁ H₃₃ N₃ O₅.H₂ O Required C 59.28H 8.29N 9.87Found C 59.30H 7.91N 9.94

EXAMPLE 104-(N-Hydroxyamino)-2R-isobutyl-3S-methylsuccinyl!-L-phenylalanine-N-(2-hydroxyethyl)amide. ##STR37##

Using the procedure described in Example 13f4-(N-Benzyloxyamino)-2R-isobutyl-3S-methylsuccinyl!-L-phenylalanine (0.4g, 0.91 mmol) was coupled with ethanolamine (61 mg, 1.00 mmol) then theproduct hydrogenated to give the title compound (51 mg, 0.13 mmol, 14%)

m.p. 208-210° C.

α!_(D) =24.0° (c=0.30, MeOH)

nu_(max) (KBr) 3280, 2960, 1635, 1540, 1450, and 1370 cm⁻¹

δ^(H) (250 MHz, D₆ -DMSO) 0.39 (3H, d, J=6 Hz, CHCH₃), 0.73 (3H, d, J=6Hz, CH(CH₃)₂), 0.80 (3H, d, J=6 Hz, CH(CH₃)₂), 1.33 (2H, t, J=10 Hz),1.94 (1H, t, J=8 Hz), 2.34 (1H, t, J=10 Hz), 2.77 (2H, t, J=12 Hz), 3.01(2H, dd, J=11.3 Hz), 3.13 (2H, q, J=6 Hz), 4.61 (1H, m), 4.66 (1H,m),7.20 (5H, m), 7.73 (1H, s), 8.18 (1H, d, J=8 Hz, NH), and 8.69 (1H, s).

δ_(C) (62.9 MHz, D₆ -DMSO) 173.5, 171.4, 138.3, 128.1, 126.3, 59.9,54.2, 46.2, 41.6, 37.4, 25.4, 24.3, 21.7, and 16.1.

Analysis calculated for C₂₀ H₃₁ N₃ O₅.0.5H₂ O Required C 59.68H 8.01N10.44 Found C 59.38H 7.71N 10.27

EXAMPLE 114-(N-Hydroxyamino)-2R-isobutyl-3S-methylsuccinyl!-L-phenylalanine-N-(3-(2-pyrrolidone)propyl)amide sodium salt. ##STR38##

4-(N-Hydroxyamino)-2R-isobutyl-3S-methylsuccinyl!-L-phenylalanine-N-(3-(1-pyrrolidone)propyl)amide (50 mg, 0.1 mmol) was dissolved in methanol (1 ml) and sodiumhydroxide solution (1.0M, 2.0 ml) added to give a homogeneous solution.The methanol was removed under reduced pressure then the residualaqueous solution freeze dried to give the title compound (53 mg, 0.1mmol, 100%).

δ_(H) (250 MHz , D₆ -DMSO) 0.44 (3H, d, J=7 Hz, CHCH₃), 0.72 (3H, d, J=6Hz, CH(CH₃)₂), 0.79 (3H, d, J=6 Hz, CH(CH₃)₂), 1.32 (2H, m), 1.53 (2H,q, J=7 Hz), 1.92 (4H, m), 2.21 (2H, t, J=8 Hz), 2.37 (1H, m), 2.79 (1H,m), 2.93 (1H, d, J=5 Hz), 3.01 (2H, m), 3.13 (2H, t, J=7 Hz), 3.26-3.43(6H, m), 4.54 (1H, m), 7.22 (5H, m), 7.85 (1H, J=7 Hz), and 8.28 (1H, d,J=8 Hz, NH).

δ_(C) (62.9 MHz , D₆ -DMSO) 174.0, 173.4, 170.8, 129.2, 127.7, 126.0,54.6, 47.0, 46.7, 37.7, 36.3, 30.7, 27.3, 25.4, 24.3, 21.5, 17.7 and16.1.

EXAMPLE 12 Collagenase inhibition activity

The potency of compounds of general formula I to act as inhibitors ofcollagenase (a metalloprotease involved in tissue degradation) wasdetermined by the procedure of Cawston and Barrett, (Anal. Biochem., 99,340-345, 1979), hereby incorporated by reference, whereby a 1 mMsolution of the inhibitor being tested or dilutions thereof wasincubated at 37° for 16 hours with collagen and collagenase (bufferedwith 25 mM Hepes, pH 7.5 containing 5 mM CaCl₂, 0.05% Brij 35 and 0.02%NaN₃). The collagen was acetylated ¹⁴ C collagen prepared by the methodof Cawston and Murphy (Methods in Enzymology, 80, 711, 1981), herebyincorporated by reference. The samples were centrifuged to sedimentundigested collagen and an aliquot of the radioactive supernatantremoved for assay on a scintillation counter as a measure of hydrolysis.The collagenase activity in the presence of 1 mM inhibitor, or adilution thereof, was compared to activity in a control devoid ofinhibitor and the results reported below as that inhibitor concentrationeffecting 50% inhibition of the collagenase (IC₅₀).

    ______________________________________    Compound of Example No.                       IC.sub.50    ______________________________________    2                  200 nM    3                  20 nM    4                  90 nM    ______________________________________

Examples of unit dosage compositions are as follows:

EXAMPLE 13

Capsules:

    ______________________________________                                Per 10,000    Ingredients       Per Capsule                                Capsules    ______________________________________    1.     Active ingredient                           40.0 mg   400 g           (Cpd of Formula I)    2.     Lactose        150.0 mg  1500 g    3.     Magnesium       4.0 mg    40 g           stearate                          194.0 mg  1940 g    ______________________________________

Procedure for capsules

Step 1. Blend ingredients No. 1 and No. 2 in a suitable blender.

Step 2. Pass blend from Step 1 through a No. 30 mesh (0.59 mm) screen.

Step 3. Place screened blend from Step 2 in a suitable blender withingredient No. 3 and blend until the mixture is lubricated.

Step 4. Fill into No. 1 hard gelatin capsule shells on a capsulemachine.

EXAMPLE 14

Tablets:

    ______________________________________                              Per 10,000    Ingredients      Per Tablet                              Tablets    ______________________________________    1.    Active ingredient                         40.0 mg  400 g          (Cpd of Form. I)    2.    Corn Starch    20.0 mg  200 g    3.    Alginic acid   20.0 mg  200 g    4.    Sodium alginate                         20.0 mg  200 g    5.    Magnesium       1.3 mg   13 g          stearate                         101.3 mg 1013 g    ______________________________________

Procedure for tablets

Step 1. Blend Ingredients No. 1, No. 2, No. 3 and No. 4 in a suitablemixer/blender.

Step 2. Add sufficient water portionwise to the blend from Step 1 withcareful mixing after each addition. Such additions of water and mixinguntil the mass is of a consistency to permit its conversion to wetgranules.

Step 3. The wet mass is converted to granules by passing it through anoscillating granulator using a No. 8 mesh (2.38 mm) screen.

Step 4. The wet granules are then dried in an oven at 140° F. (60° C.)until dry.

Step 5. The dry granules are lubricated with ingredient No. 5.

Step 6. The lubricated granules are compressed on a suitable tabletpress.

EXAMPLE 15

Intramuscular Injection:

    ______________________________________    Ingredient         Per ml. Per liter    ______________________________________    1.    Formula I compound                           10.0 mg 10 g          Active ingredient    2.    Istonic buffer   q.s.    q.s          solution pH 4.0.    ______________________________________

Procedure

Step 1. Dissolve the active ingredient in the buffer solution.

Step 2. Aseptically filter the solution from Step 1.

Step 3. The sterile solution is now aseptically filled into sterileampoules.

Step 4. The ampoules are sealed under aspetic conditions.

EXAMPLE 16

Suppositories:

    ______________________________________                               Per    Ingredients       Per Supp.                               1,000 Supp    ______________________________________    1.    Formula I compound                           40.0 mg   40 g          Active ingredient    2.    Polyethylene Glycol                          1350.0 mg                                   1,350 g          1000    3.    Polyethylene Glycol                           450.0 mg                                     450 g          4000                          1840.0 mg                                   1,840 g    ______________________________________

Procedure

Step 1. Melt ingredient No. 2 and No. 3 together and stir until uniform.

Step 2. Dissolve ingredient No. 1 in the molten mass from Step 1 andstir until uniform.

Step 3. Pour the molten mass from Step 2 into suppository moulds andchill.

Step 4. Remove the suppositories from moulds and wrap.

EXAMPLE 17

Eye Ointment

An appropriate amount of a compound of general formula I is formulatedinto an eye ointment base having the following composition:

    ______________________________________    Liquid paraffin  10%    Wool fat         10%    Yellow soft paraffin                     80%    ______________________________________

EXAMPLE 18

Topical skin ointment

An appropriate amount of a compound of general formula I is formulatedinto a topical skin ointment base having the following composition:

    ______________________________________    Emulsifying wax  30%    White soft paraffin                     50%    Liquid paraffin  20%    ______________________________________

We claim:
 1. A compound of formula I: ##STR39## wherein R¹ represents ahydrogen atom or a C₁ -C₆ alkyl, C₁ -C₆ alkenyl, phenyl, phenyl (C₁-C₆)alkyl, C₁ -C₆ alkythiomethyl, phenylthiomethyl, substitutedphenylthiomethyl, phenyl(C₁ -C₆)alkythiomethyl, pyridine-2-thiomethyl,pyridine-4-thiomethyl, thiophene-2-thiomethyl or pyrimidine-2-thiomethylgroup wherein substituted refers to up to four substituents each ofwhich independently may be C₁ -C₆ alkyl, C₁ -C₆ alkoxy, hydroxy, thiol,C₁ -C₆ alkylthiol, amino, halo, triflouromethyl, nitro, --COOH, --COONH₂or --CONHR^(A), wherein R^(A) represents a C₁ -C₆ alkyl group or theside chain of the amino acid alanine, valine, leucine, isoleucine,phenylalanine, tryptophan, methionine, glycine, serine, threonine,cysteine, tyrosine, asparagine, glutamine, aspartic acid, glutamic acid,lysine, arginine or histidine; or R¹ represents --S--R^(x) wherein R^(x)represents a group; ##STR40## R² represents a hydrogen atom or a C₁ -C₆alkyl, C₁ -C₆ alkenyl, phenyl (C₁ -C₆) alkyl, cycloalkyl (C₁ -C₆) alkyl,or cycloalkenyl (C₁ -C₆) alkyl;R³ represents an amino acid side chain ora C₁ -C₆ alkyl, benzyl, (C₁ -C₆) alkoxylbenzyl, benzyloxy (C₁ -C₆) alkylor benzyloxybenzyl group; R⁴ represents a hydrogen atom or a methylgroup; R⁵ represents a group (CH₂)_(n) A; n is an integer from 1 to 6; Arepresents a hydroxy, (C₂ -C₇)acyloxy, (C₁ -C₆)alkylthio, phenylthio, or(C₂ -C₇)acylamino; or a pharmaceutically acceptable salt or N-oxide orwhere the compound is a thio-compound, a sulphoxide or sulphone thereof.2. A compound as claimed in claim 1, in which the chiral centre adjacentthe substituent R³ has S stereochemistry.
 3. A compound as claimed inclaim 1, wherein R¹ represents a hydrogen atom or a C₁ -C₄ alkyl,phenylthiomethyl or heterocyclythiomethyl group.
 4. A compound asclaimed in claim 1, wherein R² represents a C₂ -C₆ alkyl group.
 5. Acompound as claimed in claim 1, wherein R³ represents a benzyl, 4-(C₁-C₆) alkoxyphenylmethyl or benzyloxy benzyl group.
 6. A compound asclaimed in claim 1 wherein n has the value 1, 2 or
 3. 7. A compound asclaimed in any one of claims 1 to 6, wherein R⁵ represents apyrrolidone, hydroxy, methoxy or thioethyl group.
 8. A compound selectedfrom the group consistingof:(4-(N-Hydroxyamino)-2R-isobutylsuccinyl)-L-phenylaianine-N-(2-hydroxyethyl)-amide;(4-(N-Hydroxyamino)-2R-isobutylsuccinyl)-L-phenylalanine-N-(2-hydroxyethyl)-N-methylamide;(4-(N-Hydroxyamino)-2R-isobutylsuccinyl)-L-phenylalanine-N-(2-ethylthioethyl)-amide;(4-(N-Hydroxyamino)-2R-isobutylsuccinyl)-L-phenylalanine-N-(2-N-acetylethyl)-2-aminoethylamide;(4-(N-Hydroxyamino)-2R-isobutylsuccinyl)-L-phenylalanine-N-(2-acetoxyethyl)-amide;(4(N-Hydroxyamino)-2R-isobutyl-3S-methylsuccinyl)-L-phenylalanine-N-methyl-N-(2-hydroxyethyl)-amide;(4-(N-Hydroxyamino)-2R-isobutyl-3S-methylsuccinyl)-L-phenylalanine-N-(2-hydroxyethyl)-amide;andpharmaceutically acceptable salts thereof.